Development of a highly sensitive TaqMan method based on multi-probe strategy: its application in ASFV detection

Figures

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  Figure 1.. The diagram of primers and probes for detecting B646L of ASFV. (A) Relative positions of different primers and probes. (B) The sequences of primers and probes.

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  Figure 2.. Fluorescence quantitative PCR amplification curves of different number of probes and the linear relationship. (A) Selecting representative curves of four TaqMan qPCR reactions which were based on the optimal reaction system and amplification reaction conditions. (B) The photography depicts linear relationship between average ΔCt and log2⁡n value. The coefficient of determination (R2) and equation of the regression curve (y) were calculated.

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  Figure 3.. Standard curves of the triple-probe 40/109/118 TaqMan qPCR assay. The dynamic range and detection limit were evaluated by testing 10-fold serial diluted of plasmid (from 5.0 × 106 to 5.0 × 10° copies/μl) against the Ct value. The coefficient of determination (R2) and equation of the regression curve (y) were calculated.

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  Figure 4.. Sensitivity test curves of triple-probe 40/109/118 TaqMan qPCR assay. The dynamic curves were generated by using the plasmid final concentration ranged from 5.0 × 106 to 5.0 × 10−2 copies/μl.

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  Figure 5.. Specificity of the triple-probe 40/109/118 TaqMan qPCR assay. The assay was used to amplify pUC57-B646L, PCV2, PCV3, PCV4, PEDV, PDCoV, CSFV, PRRSV, PRV. The X-axis represents the cycles, and the Y-axis represents the fluorescence data.